EVs are small (10-2000 nm) membrane-bound structures secreted by cells, and contain proteins, lipids and nucleic acids derived from their source cells. EVs can travel through biological fluids and surmount biological barriers before being taken up by their target cells, thereby transferring bioactive components and thus acting as mediators of intercellular communication. These characteristics make EV-based technologies highly exploitable for use as drug and biomarker candidates in regenerative and transplant medicine, anti-tumor therapy, and immunotherapy.
By offering extensive EV characterization services, TAmiRNA acknowledges the expanding medical value of EVs and supports EV translational and clinical research.
EV service features
The TAmiRNA one-stop shop EV characterization service extends from identification of target EV type (apoptotic bodies, exosomes, microvesicles, etc.) through EV isolation and characterization, to data analysis and reporting.
Starting from its world-leading expertise in microRNA technologies, TAmiRNA has implemented a large number of assays to study EV concentration, size, decoration with nucleic acids and proteins, as well as nucleic acid cargo. The unique EV service portfolio includes determination of absolute numbers of RNA molecules inside EVs, surface epitope staining protocols and fluorescence-based techniques for staining intracellular RNA/DNA content using NTA QUATT analysis as a means of EV manufacturing quality control.
Further knowledge is gained through parallel purification of EVs and protein complexes.
Based on this expertise, TAmiRNA has already completed small RNA sequencing analyses of more than 500 EV samples obtained from various biological samples. All services are based on best practices defined by the International Society for Extracellular Vesicles (ISEV).
Reference projects include:
- Small RNA sequencing-based characterization of EVs purified from synovial fluid, serum, plasma, urine, and conditioned media.
- SEC-purification of EVs and protein complexes from human urine samples (following 10 kDa-UF) and independent small RNA sequencing analysis of EV and protein RNA cargo
- Small RNA sequencing- and RT-qPCR-based characterization of MSC-derived EVs intended for pharmaceutical applications
- Size, concentration, and nucleic acid cargo characterization of EVs obtained from various in-vitro models and using different isolation methods
- Development of a flow-cytometry based protocol to characterize oncogenic protein surface markers on EVs.
TAmiRNA can purify EVs (and protein complexes) from biofluids, such as (platelet-poor) plasma or urine, or from cell culture supernatants, using sample volumes starting from as little as 100 µl.
Alternatively, TAmiRNA can process EV samples generated by customers that meet certain basic quality parameters.
Customers can take advantage of TAmiRNA’s specialist capabilities including:
- EV purification: By size exclusion chromatography (SEC), ultracentrifugation, or precipitation. SEC-based purification can deliver both EVs and protein complexes to study presence of RNA in both compartments in parallel.
- EV characterization: Determination of EV size, concentration, nucleic acid content (SYBR Gold), and surface epitopes (CD81, CD63, CD9) by Nanoparticle Tracking Analysis (NTA Quatt) and/or multiplex flow cytometry Assay development for the analysis of other surface epitopes on demand.
- Non-specific characterization of EV cargo: Byprotein, dsDNA, and ssRNA quantification in EV preparations. RNase/DNase/Protease treatments are used to reduce noise from surface decorations with RNA/DNA.
- exRNA analysis in EVs: Highly quantitative analysis of microRNA, mRNA, and other non-coding RNAs using robust protocols for small RNAseq, RNAseq, and RT-qPCR analysis of EVs (and protein complexes).
Click on TAmiRNA small RNA sequencing protocol for further information.
Click on Extracellular vesicles for further information.
Click on Priglinger et al. (2020) to access relevant study published at Nature.com.